Calibrator for immunoassays

Abstract

The invention generally relates to the field of immunoassays. In particular, the invention relates to use of a calibrator material to calibrate immunoassays for autoantibodies.

1 .- 36 . (canceled) 
     
     
         37 . A method of calibrating a GBU4-5-specific immunoassay for detecting an autoantibody in a human serum sample, wherein the method comprises:
 (a) providing a calibration material comprising an antibody immunologically specific for a tag that is a component of a tagged GBU4-5 antigen;   (b) contacting serial dilutions of the calibration material with a static amount of a control protein comprising the tag such that the antibody specific for the tag binds to the control protein comprising the tag in each of the serial dilutions of the calibration material;   (c) contacting serial dilutions of the human serum sample with a static amount of the tagged GBU4-5 antigen such that the autoantibodies in each of the serial dilutions of the human serum sample bind to the tagged GBU4-5 antigen.   (d) measuring amounts of specific binding between the antibody specific for the tag and the control protein comprising the tag present in each of the serial dilutions of the calibration material and amounts of specific binding between the autoantibodies in the human serum sample and the tagged GBU4-5 antigen concurrently on a single assay plate;   (e) plotting or calculating a curve of the amount of the specific binding versus the dilution of the calibration material for each dilution of calibration material used in step (b); and   (f) extrapolating, from the curve, an equivalent dilution of the calibration material for a corresponding measure of specific binding between the autoantibodies in the human serum sample and the tagged GBU4-5 antigen, to thereby calibrate the human serum sample and monitor run-to-run variation in the immunoassay.   
     
     
         38 . The method of  claim 37 , wherein the antigen of part (c) is used at a concentration greater than 20 nM. 
     
     
         39 . The method of  claim 38 , wherein the antigen of part (c) is used at a concentration in the range of from 20 nM to 180 nM. 
     
     
         40 . The method of  claim 38 , wherein the antigen of part (c) is used at a concentration in the range of from 50 nM to 160 nM. 
     
     
         41 . The method of  claim 37 , wherein the tag is a protein or peptide tag. 
     
     
         42 . The method of  claim 41 , wherein the protein or peptide tag is a histidine tag or biotin tag. 
     
     
         43 . The method of  claim 37 , wherein the amounts of specific binding between the antigen and autoantibody are detected using a colorimetric, chemiluminescent or fluorescent system. 
     
     
         44 . The method of  claim 37 , wherein the amounts of specific binding between the antigen and autoantibody are detected using a labelled secondary anti-human immunoglobulin antibody. 
     
     
         45 . The method of  claim 44 , wherein the labelled secondary anti-human immunoglobulin antibody is anti-IgG or anti-IgM. 
     
     
         46 . The method of  claim 37 , wherein the autoantibody is of IgG or IgM isotype. 
     
     
         47 . The method of  claim 37 , wherein the antigen is a recombinant tumor marker antigen. 
     
     
         48 . The method of  claim 37 , wherein the curve of the amount of the specific binding versus the dilution of the calibration material is a four parameter logistic plot. 
     
     
         49 . The method of  claim 37 , wherein the antibody immunologically specific for the tag is a human antibody. 
     
     
         50 . The method of  claim 37 , wherein the antibody immunologically specific for the tag is a native autoantibodies or a recombinant antibody. 
     
     
         51 . The method of  claim 37 , wherein the control protein is a recombinant peptide comprising a histidine tag. 
     
     
         52 . The method of  claim 37 , wherein the human serum sample is obtained from a subject with a pulmonary nodule. 
     
     
         53 . The method of  claim 37 , wherein the method is used to calibrate a GBU4-5-specific immunoassay for diagnosis of lung cancer. 
     
     
         54 . A method of assessing the effectiveness of an anti-cancer treatment of lung cancer in a subject, comprising:
 (a) obtaining a biological sample from the subject that is undergoing the anti-cancer treatment;   (b) measuring the amount of autoantibodies specific to the tumor antigen GBU4-5 in the biological sample;   (c) comparing the amount of the autoantibodies in the biological sample to a pre-determined clinical threshold that indicates that the subject is likely to have lung cancer; and   (d) determining, based on the step (c), whether the anti-cancer treatment is effective, wherein a higher amount of the autoantibodies in the biological sample than the pre-determined clinical threshold indicates the anti-cancer is ineffective while a lower amount of the autoantibodies in the biological sample than the pre-determined clinical threshold indicates the anti-cancer is effective.   
     
     
         55 . The method of  claim 54 , wherein the subject receives a different anti-cancer treatment if the amount of autoantibodies in the biological sample is higher than the pre-determined clinical threshold. 
     
     
         56 . The method of  claim 54 , further comprising step (e): continuing to administer the same anti-cancer treatment to the subject if the amount of autoantibodies in the biological sample is lower than the pre-determined clinical threshold.